Distinctive Lacritin Cleavage-Potentiated Bactericidal Alteration of the P. aeruginosa Transcriptome

Abstract

Lacritin is a tear, saliva, plasma and cerebrospinal fluid glycoprotein with broad polypharmacology. Selective deficiency of its bioactive monomeric form appears to be deleterious for ocular surface health for which replacement therapy is beneficial. Its cleavage-potentiated C-terminus represented by the N-104 proteoform in tears is bactericidal and synergizes with the tear thrombin peptide GKY20. In the pathogenic and multidrug resistant PA14 strain of P. aeruginosa, we recently discovered that N-104 binds to the outer-membrane lipoprotein YaiW to gain access to the periplasm where it targets and inhibits the inner-membrane ferrous iron transporter FeoB (of FeoABC) as well as PotH, a subunit of the polyamine transporter PotFGHI. Further, PA14 gene expression shifts toward anaerobic respiratory pathways. Here we explore N-104-associated transcriptional changes over a broader range of functional categories pointing to a reduction in: (i) virulence by suppressed gene expression of virulence factors AprA and LasA; and Hcp1 and PsrA necessary for the respective assembly of type VI and III secretion systems, (ii) fitness (less AtsC, MgtA), (iii) metabolism (less AdhA, AtsC, GcvH2, GcvP2, FadE1, SsuD, SsuF, TauB, TauD, UspK, UspN), (iv) stress response (less UnG, PfpI, RmF), (v) proteostasis (less ClpB, GrpE, HtpG), (vii) quorum sensing (less CifR, GcvH2, GcvP2, PsrA, QuiP), and (viii) survival under anaerobic conditions (less AdhA, MhR, ModA, UspKLNO). Upregulated genes are directed towards enhancing PA14: (i) multidrug (more OprJ of MexCD-OprJ) and (ii) tellurite (more TerC) efflux, coupled with a seemingly PA14 survival attempt at (iii) anaerobic respiration (more NosR), (iv) translational fidelity (more QueE, RimP, TrmD) and (v) metabolism (CysT, MoaA1, Sbp, SsuA, SsuE). The overlap with aminoglycosides (4.3%), beta-lactams (0%), cyclic peptides (2.5%), fluoroquinilones (0%) and macrolide (1.9%) classes of antibiotics in P. aeruginosa was minimal. Thus, N-104 appears to widely perturb PA14 fundamental processes in a distinctive manner.